Vitamin D attenuates cerebral artery remodeling through VDR/AMPK/eNOS dimer phosphorylation pathway after subarachnoid hemorrhage in rats.

The role of vitamin D3 (VitD3) in the upregulation of osteopontin (OPN) and eNOS in the endothelium of cerebral arteries after subarachnoid hemorrhage (SAH) is investigated. The endovascular perforation SAH model in Sprague-Dawley rats ( n = 103) was used. The VitD3 pretreatment (30 ng/kg) increased endogenous OPN and eNOS expression in cerebral arteries compared with naïve rats ( n = 5 per group). Neurobehavioral scores were significantly improved in Pre-SAH+VitD3 group compared with the SAH group. The effects of VitD3 were attenuated by intracerebroventricular (i.c.v) injections of siRNA for the vitamin D receptor (VDR) and OPN in Pre-SAH+VitD3+VDR siRNA and Pre-SAH+VitD3+OPN siRNA rats, respectively ( n = 5 per group). The significant increase of VDR, OPN and decrease of C44 splicing in the cerebral arteries of Pre-SAH+VitD3 rats lead to an increase in basilar artery lumen. The increase in VDR expression led to an upregulation and phosphorylation of AMPK and eNOS, especially dimer form, in endothelium of cerebral artery. The results provide that VitD3 pretreatment attenuates cerebral artery remodeling and vasospasm through the upregulation of OPN and phosphorylation of AMPK (p-AMPK) and eNOS (p-eNOS) at Ser1177-Dimer in the cerebral arteries. Vitamin D may be a useful new preventive and therapeutic strategy against cerebral artery remodeling in stroke patients.

Treatment strategies of spinal arteriovenous fistulas and malformations: timing matters.

Spinal vascular malformations are a rare heterogeneous entity of spinal lesions with abnormal blood vessels. They may occur extra- and intradurally, peri- and intramedullary, with confined or complex configuration, and at various levels of the spine. Associated clinical symptoms are extremely variable and may range from diffuse pain, to progressive myelopathy, and even to acute hemorrhage with severe neurological deficits. Often, the patients present with significant spinal cord injury before the spinal arteriovenous lesions are diagnosed. Early recognition of the pathology is essential to cease potential disease progression and neurological deterioration. Therefore, the purpose of this review was to describe the current findings in spinal arteriovenous lesions focusing on the timing of treatment and treatment modality.

Elucidating the prognostic significance of KRAS, NRAS, BRAF and PIK3CA mutations in Chinese patients with metastatic colorectal cancer.

AIM: The prognostic significance of KRAS, NRAS, PIK3CA and BRAF mutations was evaluated in Chinese patients with metastatic colorectal cancer (CRC). METHOD: Tumor samples from 183 patients were retrospectively tested for KRAS, NRAS, PIK3CA and BRAF mutations. Multivariate analysis was performed to determine the relationship between mutational status, drug response and survival. RESULT: Over 70% of patients received two or more lines of chemotherapy, 50% had cetuximab and 18% had bevacizumab. The prevalence of KRAS, NRAS, BRAF and PIK3CA mutations was 45, 3.2, 5 and 20%, respectively. For the entire cohort, the median overall survival was 24 months (95% confidence interval [CI] = 20.4-26.4 months). Of the genes tested, only KRAS mutation was an independent prognostic factor with a multivariate hazard ratio of 1.5 (95% CI = 1.05-2.16, P = 0.03). In the subgroup of patients who received cetuximab-based therapy in the first-line setting, KRAS mutation was associated with a lack of response to chemotherapy (28% vs 66%, chi-square, P = 0.01). Patients with KRAS mutant tumors (or KRAS wild-type tumors that harbored BRAF and/or PIK3CA mutations) tended to have lower response rates to chemotherapy and/or cetuximab (P = not significant). The number of NRAS mutant cases was too small to allow any statistical analysis. CONCLUSION: The prevalence of KRAS, NRAS, BRAF and PIK3CA mutations in this cohort is consistent with reports from non-Asian populations, and KRAS mutation has both prognostic and predictive significance in Chinese patients with metastatic CRC.

Intermittent versus continuous erlotinib with concomitant modified "XELOX" (q3W) in first-line treatment of metastatic colorectal cancer: correlation with serum amphiregulin and transforming growth factor alpha.

BACKGROUND: This study evaluated the activity of 2 schedules of erlotinib in combination with chemotherapy, and the prognostic significance of serum amphiregulin (AREG) and transforming growth factor alpha (TGFa) in metastatic colorectal cancer. METHODS: A total of 60 untreated patients were randomized to a "continuous" (CON; erlotinib 100 mg daily) or an "intermittent" (INT; erlotinib 150 mg on alternate day on day 2 to 14, then 150 mg daily on days 15 to 21) schedule of erlotinib with a modified XELOX (capecitabine plus oxaliplatin) regimen. Serum levels of AREG and TGFa were determined serially. RESULTS: Baseline characteristics were similar between the 2 arms. Of the 58 patients evaluated for response, there was a nonsignificant trend toward a slightly higher overall response rate in the INT arm (66.7%) versus the CON arm (56.7%). At a median follow-up of 2.8 years, the median overall survival was 18.8 months (95% confidence interval = 11.3-22.9 months) and 20.7 months (95% confidence interval = 12.5-31 months, P = .19) for the CON and INT arm, respectively. KRAS mutation did not predict drug response. The 2 arms did not differ significantly in toxicity. Baseline serum TGFa was an independent predictor of progression-free survival, whereas a drop in serum TGFa and AREG levels following 3 to 4 cycles of treatment were associated with shorter progression-free survival and overall survival, respectively. CONCLUSIONS: The intermittent erlotinib schedule was associated with a higher response rate, although this is not statistically significant. Serum TGFa and AREG levels have prognostic significance in erlotinib-treated patients with colorectal cancer, and further studies are warranted.

Supportive care needs in Hong Kong Chinese women confronting advanced breast cancer.

BACKGROUND: Women with advanced breast cancer (ABC) are living longer, so understanding their needs becomes important. This cross-sectional study investigated the type and extent of unmet supportive care needs in Hong Kong Chinese women with advanced breast cancer. METHODS: Face-to-face interviews were conducted among women with stage III or stage IV disease mostly awaiting chemotherapy (76%) to identify unmet needs using the Supportive Care Needs Survey Short Form, psychological morbidity using the Hospital Anxiety and Depression Scale, symptom distress using the Memorial Symptom Assessment Scale, and satisfaction with care using the Patient satisfaction questionnaire (PSQ-9). RESULTS: About 27-72% of 198/220 (90%) women (mean age = 53.4 ± 9.74 (standard deviation) years) identified needs from the health system, information, and patient support (HSIPS) domain as the top 15 most prevalent unmet needs. 'having one member of hospital staff with whom you can talk to about all aspect of your condition, treatment, and follow-up' was most cited by 72% of the patients, with remaining unmet needs addressing mostly desire for information. Unmet need strength did not differ between women with stage III and stage IV disease, whereas women with first time diagnosis reported greater health system and information unmet needs compared with women with recurrent disease. Stepwise multiple regression analyses revealed that symptom distress was consistently positively associated with all but sexuality need domains, whereas low satisfaction with care was associated with HSIPS (ß = 3.270, p < 0.001) and physical and daily living (ß = 2.810, p < 0.01) domains. DISCUSSIONS: Chinese women with ABC expressed need for continuity of care and improved information provision. High symptom distress was associated with lower levels of satisfaction with care. These unmet needs appear to reflect current care services shortcomings.

Comparison between ambulatory infusion mode and inpatient infusion mode from the perspective of quality of life among colorectal cancer patients receiving chemotherapy.

Folfox and Folfiri are active chemotherapy treatments used in advanced colorectal cancer. The total admission for these treatments has been significantly increasing in the study hospital. An ambulatory infusion programme was launched to address the problems of long waiting list and bed shortage. The study objective was to compare the quality of life between patients receiving the chemotherapy treatments in the ambulatory infusion group and the inpatient infusion group. The results showed that some demographic variables of education level, family role and employment status were determinants of the treatment mode. Patients in the ambulatory infusion group of social and global domains performed better than the inpatient group. However, the inpatient group had better quality of life of nauseated symptom at different stages. Some nursing educations should be reinforced to address the symptom management for patients receiving ambulatory infusion at home. The ambulatory infusion programme offered considerable quality of life benefits to colorectal cancer patients receiving chemotherapy.

New utility of an old marker: serial alpha-fetoprotein measurement in predicting radiologic response and survival of patients with hepatocellular carcinoma undergoing systemic chemotherapy.

PURPOSE: There are limitations in using radiologic evaluation to assess the treatment outcome of patients with hepatocellular carcinoma (HCC). The use of serial alpha-fetoprotein (AFP) in monitoring response has not been rigorously evaluated. We aimed to study the clinical value of AFP trend in an attempt to validate AFP as a surrogate serologic end point. PATIENTS AND METHODS: Participants from a phase III randomized trial of systemic chemotherapy in HCC were studied. Serum AFP was prospectively collected in parallel with clinical and radiologic outcome. AFP response was defined as a decrease in AFP of more than 20% after a minimum of two cycles of chemotherapy. We studied the relationship between AFP response and treatment outcome in terms of radiologic response and overall survival. RESULTS: Of 188 patients, 117 patients with elevated serum AFP (> 20 microg/L) and documented radiologic evaluation had received at least two cycles of chemotherapy. A total of 47 AFP responders were identified. AFP responders had better survival than nonresponders (13.5 v 5.6 months, respectively; P < .0001), and AFP response was strongly associated with radiologic response (P < .0001). Multivariate analysis suggested that AFP response was significantly associated with survival (hazard ratio, 0.413; 95% CI, 0.273 to 0.626; P < .0001). AFP responses were frequently observed in patients with radiologically stable disease (SD) and tended to identify a subgroup of SD patients with better survival. CONCLUSION: Serial AFP measurement is useful in prognostication and monitoring treatment response in HCC patients undergoing systemic chemotherapy. Incorporation of AFP response into the criteria evaluating treatment outcome should be considered in clinical practice and clinical trials of novel agents in HCC.

Hepatitis B viral load predicts survival of HCC patients undergoing systemic chemotherapy.

UNLABELLED: HCC is a common cause of morbidity and mortality. For patients who are not candidates for curative surgery, systemic chemotherapy is one of the standard treatments. In parts of China and the Far East, over 80% of HCC patients have chronic HBV infection. In this study, we aimed to assess the relationship between pre-chemotherapy HBV viral load and the survival of HCC patients. HBV infection status was determined prior to chemotherapy in 188 patients, 170 of whom had evidence of HBV chronic infection/exposure (160 hepatitis B surface antigen [HBsAg]-positive, 10 HBsAg-negative/hepatitis B core antibody-positive). Of these, 125 had pretreatment HBV DNA levels determined via real-time PCR. Virological data were analyzed using conventional clinical variables to identify factors that influenced survival. Multivariate analysis revealed that high total bilirubin (P = 0.0016; hazard ratio = 1.040 per 1 muM increase; 95% CI 1.015-1.065), HCV infection (P = 0.0095; hazard ratio = 6.955; 95% CI 1.606-30.129), and high HBV DNA level (P = 0.0217; hazard ratio = 1.650; 95% CI 1.076-2.531) affected survival significantly. Exploratory analysis revealed that high levels of pretreatment HBV DNA had a significantly higher incidence of severe hepatitis during chemotherapy. CONCLUSION: For HCC patients with HBV chronic infection/exposure, a high viral load prior to treatment is an adverse factor for survival and may be associated with a higher incidence of severe hepatitis during chemotherapy. Future strategies to improve the prognosis of HCC patients undergoing chemotherapy should consider supportive therapy that incorporates antiviral therapies to reduce HBV viral load.

A randomized phase III study of doxorubicin versus cisplatin/interferon alpha-2b/doxorubicin/fluorouracil (PIAF) combination chemotherapy for unresectable hepatocellular carcinoma.

BACKGROUND: Single-agent doxorubicin has been widely used to treat unresectable hepatocellular carcinoma (HCC), but the response rate is low (< 20%) and there is no convincing evidence for improved survival. Cisplatin, interferon, doxorubicin, and fluorouracil (PIAF) used in combination, by contrast, has shown promise in a phase II study. We compared doxorubicin to PIAF in patients with unresectable HCC in a phase III trial. METHODS: Patients with histologically confirmed unresectable HCC were randomly assigned to receive either doxorubicin or PIAF every 3 weeks, for up to six cycles. The primary endpoint was overall survival, and secondary endpoints were response rate and toxicity. Survival differences were calculated using the Kaplan-Meier method. Treatment groups were compared for differences in the incidence of adverse events using chi-square tests. All statistical tests were two-sided. RESULTS: The median survival of the doxorubicin and PIAF groups was 6.83 months (95% confidence [CI] = 4.80 to 9.56) and 8.67 months (95% CI = 6.36 to 12.00), respectively (P = 0.83). The hazard ratio for death from any cause in the PIAF compared with the doxorubicin groups was 0.97 (95% CI = 0.71 to 1.32). Eighty-six of the 94 patients receiving doxorubicin and 91 of the 94 receiving PIAF were assessable for response. The overall response rates in the doxorubicin and PIAF groups were 10.5% (95% CI = 3.9% to 16.9%) and 20.9% (95% CI = 12.5% to 29.2%), respectively. Neutropenia, thrombocytopenia, and hypokalemia were statistically significantly more common in patients treated with PIAF than in patients treated with doxorubicin. CONCLUSION: Although patients on PIAF had a higher overall response rate and better survival than patients on doxorubicin, the differences were not statistically significant. PIAF was also associated with increased treatment-related toxicity. The prognosis of patients with unresectable HCC remains poor.

Liver resection after irinotecan, 5-fluorouracil, and folinic acid for patients with unresectable colorectal liver metastases: a multicenter phase II study by the Cancer Therapeutic Research Group.

The main objectives of this study were to assess the use of irinotecan, 5-fluorouracil (5-FU), and leucovorin (FA) as neoadjuvant chemotherapy for patients with unresectable colorectal liver metastases and to determine the response rate and proportion of patients that could be down-staged to resectable tumors. Forty patients were treated with irinotecan (180 mg/m2 over 30 min) on d 1, FA (200 mg/m2 over 30 min) followed by 5-FU (400 mg/m2 bolus and continuous infusion of 600 mg/m2 over 22 h) on d 1 and 2 every 2 wk. The overall response rate was 55% (95% CI: 39.5-70.4%). The progression-free survival was 12.1 mo (95% CI: 11.4-14.8 mo). The median overall survival was 20 mo (95% CI: 17.7-26.6 mo). Four patients (10%) have undergone liver resection after a median of eight cycles. Those patients remained alive with a median follow up period of 33 mo. The principal grade 3-4 toxicity was neutropenia in 20 patients (50%). We conclude that the regimen of irinotecan/5-FU/FA was highly active in patients with colorectal cancer and liver metastases with limited toxicity. In a subgroup of patients with initial inoperable liver metastases, this regimen was able to down-stage the disease to an operable stage.

Prevention of hepatitis B virus reactivation in patients with nasopharyngeal carcinoma with lamivudine.

PURPOSE: Although the mainstay of treatment of patients with nasopharyngeal carcinoma (NPC) had been radiotherapy, chemotherapy has increasingly been adopted in conjunction with radiation and in advanced disease. In parts of Asia where NPC is prevalent, it is also known that around 10% of the population has chronic hepatitis B virus (HBV) infection. Cancer patients who are HBV carriers are frequently complicated by HBV reactivation during chemotherapy. This may result in liver damage, which disrupts anticancer therapy and compromises the patients' prognosis. In its most severe form, fatal hepatic failure may occur. With the increasing use of chemotherapy in NPC, the occurrence of HBV reactivation is likely to increase further. Although recent reports have suggested that the antiviral agent lamivudine may reduce HBV reactivation and its associated morbidity, there has been no data on this aspect in NPC patients. This study assessed the role of lamivudine in preventing HBV reactivation and its associated morbidity in NPC patients who have chronic HBV infection and are undergoing chemotherapy. MATERIALS AND METHODS: Two groups were studied. One group consisted of 16 patients who received prophylactic lamivudine prior to and until 8 weeks after discontinuing chemotherapy. The other comprised 21 historical control subjects who underwent chemotherapy without prophylactic lamivudine. The outcomes were compared. RESULTS: With prophylactic lamivudine, there were significantly fewer incidences of hepatitis (6.7% vs 33.3%, P = 0.047) and HBV reactivation (0% vs 28.6%, P = 0.027), and less disruption of chemotherapy (18.8% vs 67.7%, P = 0.045). CONCLUSION: Prophylactic lamivudine significantly reduces the incidence and morbidity of HBV reactivation in NPC patients undergoing chemotherapy.

Use of lamivudine to prevent hepatitis B virus reactivation during chemotherapy in breast cancer patients.

In parts of Asia, about 10% of the population have chronic hepatitis B virus (HBV) infection, and cancer patients who are HBV carriers are frequently complicated by HBV reactivation while receiving cytotoxic chemotherapy. The condition may result in varying degrees of liver damage, causing disruption in chemotherapy and compromising the patients' prognosis. With the increasing use of chemotherapy paralleling the rise in breast cancer incidence, the occurrence of HBV reactivation is likely to further increase. Recent reports have suggested that the anti-viral agent, lamivudine, may reduce HBV reactivation and its associated morbidity. However, most studies are based on small series of lymphoma patients, while information on the other high risk population, namely breast cancer patients, has been lacking. In this study, we studied the role of lamivudine in preventing HBV reactivation and its associated morbidity in breast cancer patients with chronic HBV infection who were planned for chemotherapy. Two groups were studied. One group consisted of 31 patients who received 'prophylactic lamivudine' prior to and until 8 weeks after discontinuing chemotherapy. The other comprised of 61 historical controls who underwent chemotherapy without prophylactic lamivudine. The outcomes, in terms of the efficacy of lamivudine in reducing the incidence of HBV reactivation, and diminishing morbidity during chemotherapy were compared. The results revealed that in the prophylactic lamivudine group, despite a significantly higher proportion receiving anthracyclines, there was significantly fewer incidences of hepatitis (12.9 vs. 59.0%, p < 0.001), less HBV reactivation (6.5. vs. 31.1%, p=0.008), and less disruption of chemotherapy (16.1% vs. 45.9%, p=0.006). We conclude that prophylactic lamivudine significantly reduces the incidence of HBV reactivation and the overall morbidity of breast cancer patients undergoing chemotherapy.

Lamivudine for the prevention of hepatitis B virus reactivation in hepatitis B s-antigen seropositive cancer patients undergoing cytotoxic chemotherapy.

PURPOSE: For cancer patients receiving cytotoxic chemotherapy, hepatitis B virus (HBV) reactivation is a well described complication resulting in varying degrees of liver damage. The objectives of this study were to assess the efficacy of the antiviral agent lamivudine in reducing the incidence of HBV reactivation and diminishing morbidity and mortality of cancer patients with chronic HBV infection during chemotherapy. PATIENTS AND METHODS: Two groups were compared in this nonrandomized study. The prophylactic lamivudine group consisted of 65 patients in a phase II study who were treated with lamivudine before and until 8 weeks after discontinuing chemotherapy. The historical controls consisted of 193 consecutive patients who underwent chemotherapy without prophylactic lamivudine. Significant prognosticators for the development of HBV reactivation were determined based on data from the controls. Potential confounding factors were identified between the two groups. The outcomes were compared. RESULTS: In the controls, lymphoma and anthracycline usage were factors identified to be associated with reactivation. The two groups were comparable in most baseline characteristics, although in the prophylactic lamivudine group, there were significantly more patients with lymphoma and receiving anthracyclines. In the prophylactic lamivudine group, there was significantly less HBV reactivation (4.6% v 24.4% in the controls; P <.001), fewer incidences of hepatitis (17.5% v 44.6%; P <.0001) that were less severe (4.8% v 18.7%; P =.0005), and less disruption of chemotherapy (15.4% v 34.6%; P =.0029). The reduction in overall mortality was not statistically different. CONCLUSION: Prophylactic lamivudine significantly reduced the incidence of HBV reactivation and the overall morbidity of cancer patients undergoing chemotherapy.

Hepatitis B virus reactivation in breast cancer patients receiving cytotoxic chemotherapy: a prospective study.

Breast cancer is a rapidly increasing problem in many developing countries, and cytotoxic chemotherapy is now an integral part of its management. In several developing countries, the carriage of hepatitis B virus (HBV) in cancer patients may be as high as 12%, and such patients are at risk of developing fatal HBV reactivation during chemotherapy. HBV reactivation is well recognized in patients with hematological malignancies, but limited data are available on patients with other, more common, cancers, such as breast cancer. Recent data have suggested that increased viral replication, an indication of HBV reactivation, may precede clinical hepatitis. In the absence of serial HBV DNA monitoring, HBV reactivation during chemotherapy may have been underestimated. In this prospective study, breast cancer patients who were hepatitis B surface antigen (HBsAg) seropositive were followed up during chemotherapy. The main objectives were to determine the incidence of HBV reactivation in breast cancer patients undergoing conventional chemotherapy; to investigate whether "serial HBV DNA monitoring" improves the accuracy of diagnosing HBV reactivation when compared with previous schema that only measured HBV DNA at the time of clinical hepatitis ("conventional monitoring"); and to assess the clinical consequences as a result of developing the condition. The secondary objective was to identify risk factors associated with this condition. Over an 18-month period, 41 patients were studied. Ten developed HBV reactivation by conventional monitoring criteria, but with serial HBV DNA monitoring, seven additional patients were diagnosed when increased HBV DNA levels were detected before, but not concomitant with, clinical hepatitis. Thus, a total of 17 patients (41%) developed HBV reactivation. Premature termination of chemotherapy or delay in treatment schedules occurred in 71% of the patients who developed viral reactivation, as compared with 33% in those who did not develop the condition (P = 0.019). No risk factors associated with the development of HBV reactivation could be identified. Serial monitoring of HBV DNA, in addition to liver function, increases the sensitivity of diagnosing of HBV reactivation, and helps explain some cases that would otherwise be labeled as "cryptogenic hepatitis," for which concomitant HBV DNA measured at the time of hepatitis was undetectable. The present study highlights the importance of monitoring HBsAg-seropositive patients who are receiving chemotherapy for common solid tumors such as breast cancer.

Phase II study of docetaxel and epirubicin in Chinese patients with metastatic breast cancer.

The efficacy and safety of docetaxel-epirubicin chemotherapy in the treatment of metastatic breast cancer was investigated in Chinese women. Three-weekly cycles comprised epirubicin 75 mg/m2 i.v. followed 1 h later by docetaxel 75 mg/m2 i.v. After 3 cycles, responding patients received a further 3 cycles, followed by 3 cycles of docetaxel alone. Forty-six patients entered the study, of whom 37% had received prior adjuvant chemotherapy. Three patients withdrew due to toxicity and were not evaluable for response. There were five complete responses and 31 partial responses, giving an overall response rate of 83.7% (95% CI 72.7-94.8%). The median time to progression was 10.96 months (95% CI 7.76-12.86) and median survival was 24.2 months (95% CI 16.6-). The most common grade 3/4 adverse events were neutropenia (96% of patients) and neutropenia with fever (39%). Hepatotoxicity occurred in six patients, two being attributable to hepatitis B virus reactivation. No patients suffered grade 3/4 cardiac toxicity and there were no treatment-related mortalities. Quality of life aspects deteriorated after 3 cycles, but there was a trend towards improved emotional aspects after 9 cycles. We conclude that docetaxel-epirubicin chemotherapy is highly effective for recurrent metastatic/locoregional breast cancer, with myelosuppression being the main toxicity.

Acute toxicity of adjuvant doxorubicin and cyclophosphamide for early breast cancer -- a retrospective review of Chinese patients and comparison with an historic Western series.

A cohort of 85 Chinese breast cancer patients who received adjuvant chemotherapy with doxorubicin and cyclophosphamide was found to have a significantly higher incidence of grade 3 (n=44, 52%) and grade 4 (n=21, 25%) neutropenia when compared with an historic Western cohort. Also noted was a higher incidence of hepatotoxicity (n=8, 9%). When compared to Caucasian patients, the higher myelotoxicity in our patients may be related to ethnic variation in susceptibility to chemotherapy-related toxicity, lower body mass index with higher percentage of body fat composition, and the popular practice of concurrent alternative medicine during chemotherapy. The higher incidence of hepatoxicity was possibly associated with endemic chronic hepatitis B infection in this geographical area.

Frequent detection of the replicative form of TT virus DNA in peripheral blood mononuclear cells and bone marrow cells in cancer patients.

The TT virus (TTV), a member of a family of human viruses related to the circoviridae viruses, was associated initially with acute and chronic liver diseases. TTV consists of a single-stranded, circular DNA genome of 3.8 kilobases (kb) and at least three open reading frames (ORFs). The objective of the present study was to determine whether or not TTV replicated in peripheral blood mononuclear cells (PBMCs) and bone marrow cells (BMCs). DNA was extracted from the PBMCs or BMCs of 153 cancer patients and from the PBMCs of 50 healthy blood donors (the controls). By using a single round of polymerase chain reaction (PCR), TTV was detected in 98.6% (141 of 143) of the PBMCs and in 90% (9 of 10) of the BMCs from cancer patients. TTV DNA was detected in significantly fewer control subjects at 86% (43 of 50; P < 0.05). Strand-specific PCR (SSPCR) targeting the ORF2 of the common genotypes of TTV was developed specifically to detect TTV positive or negative strand DNA and to examine TTV replication. TTV positive strand DNA, which may be an intermediate of viral replication, was detected in 55.3% (78 of 141) of the TTV-infected PBMCs of the cancer patients and in 7% (3 of 43) of the controls (P < 0.001). The replicative form of TTV was also detectable in 55.6% (5 of 9) of the TTV-infected BMCs. The existence of double-strand (positive and negative strands) TTV DNA in PBMCs and BMCs of the cancer patients was also supported by the finding that TTV DNA extracted from these cells was resistant to S1 nuclease. Using in situ hybridization, TTV DNA was also demonstrated to be present in the nucleus of PBMCs. It is concluded that replicative intermediate forms of TTV DNA are present in both PBMCs and BMCs, indicating that blood cells may be a site of TTV replication.